J. Phys. Chem. A 2002, 106, 3235-3242
3235
Aqueous Complexation Equilibria of meso-Tetrakis(4-carboxyphenyl)porphyrin with Viologens: Evidence for 1:1 and 1:2 Complexes and Induced Porphyrin Dimerization Suzanne E. Clarke, Carl C. Wamser,* and Harry E. Bell Department of Chemistry, Portland State UniVersity, Portland, Oregon 97207-0751 ReceiVed: August 30, 2001; In Final Form: December 20, 2001
UV-vis spectra of 5,10,15,20-tetrakis(4-carboxyphenyl)porphyrin (TCPP) have been studied in dilute (1.0 µM) aqueous solution at pH 7.0 as a function of added electrolytes, including phosphate buffer, methyl viologen dichloride (MV), and propyl viologen sulfonate (PVS). At concentrations e1.0 µM and pH g 7.0, TCPP is considered to be tetraanionic and unaggregated. It shows a strong B (Soret) band at 414.2 nm that can be decomposed into three Gaussians, and follows Beer’s law with ) 4.8 × 105 M-1 cm-1 at pH 7.0 in 5.0 mM phosphate buffer. Deviations from Beer’s law appear above 2 µM under the above conditions; linear regions for Beer’s law plots depend on the pH, temperature, and ionic strength. Even around 1 µM, the Soret absorption strength is a very sensitive function of electrolyte concentration; for example, ) 3.6 × 105 M-1 cm-1 at pH 7.0 in 62 mM phosphate buffer, and ) 5.1 × 105 M-1 cm-1 in aqueous solution of minimal ionic strength (0.1 mM NaOH, pH 10). In the titration of TCPP with MV, two types of experiments were performed, at low ionic strength (which necessarily increased in the later stages of the titration) or at high (effectively constant) ionic strength. In 5.0 mM phosphate buffer at pH 7.0 two isosbestic points are observed, at 418.4 nm (in the range of 40-300 µM MV) and at 421.5 nm (15-90 mM MV), considered to represent the formation of 1:1 and 1:2 complexes (porphyrin-viologen). By nonlinear regression analysis, the complexation constants are calculated to be K1 ) 3350 M-1 and K2 ) 68 M-1. Application of principal component analysis identifies the spectra of both complexes and gives K1 ) 2200 M-1 and K2 ) 100 M-1. The complexation constants, the spectra of the complexes, and the positions of the isosbestic points vary with the buffer concentration. At 62 mM phosphate buffer, K1 ) 600 M-1 and K2 ) 40 M-1 by nonlinear regression analysis (K1 ) 1200 M-1 and K2 ) 140 M-1 by principal component analysis). 1H NMR spectra indicate that both TCPP and MV undergo concentration-dependent resonance shifts that can be correlated with the calculated complexation constants. Similar titration of TCPP with PVS also shows evidence for 1:1 and 1:2 complexes, but with lower equilibrium constants compared to those from titration with MV. At high PVS concentrations (>25 mM) a blue-shifted Soret band attributed to TCPP dimer is observed. The same spectrum is observed under a variety of other conditions, including the presence of a cationic surfactant (4 µM CTAB), very high ionic strength (3 M NaCl), or a polycationic complexing agent. These effects illustrate the variety of influences that affect the porphyrin absorption spectrum in aqueous solution and offer cautions about the conditions of experimental control and data analysis that are necessary to extract meaningful titration data.
Introduction Porphyrins are frequently prepared with ionic substituents to confer water solubility; the special properties of such porphyrins have been thoroughly reviewed recently.1 In particular, ionic porphyrins are commonly used as water-soluble electron-transfer photosensitizers,2 and are often used in conjunction with viologens (bipyridinium dications) as electron acceptors.3 Porphyrin-viologen pairs have been used for photoinduced electron transfer in a variety of solution-phase and supramolecular assemblies.4,5 Numerous complications can arise from even this simple pairing of functional units. For example, it has been wellestablished that porphyrins have a tendency to form homoaggregates in aqueous solution.1,6-13 Porphyrins of opposite charge type show a strong tendency to form heterodimers or aggregates.1,14-18 In addition, porphyrins undergo complexation with charged species ranging from DNA to metal ions.1 Complexation with viologens is most pronounced with anionic por* To whom correspondence should be addressed. E-mail: wamserc@ pdx.edu.
phyrins,19-33 but neutral porphyrins can also complex viologens under appropriate circumstances.34,35 Furthermore, solvent and other environmental influences exert significant effects on porphyrin spectroscopy.36 In this paper, we characterize in detail the behavior of a dilute aqueous solution of an anionic free base porphyrin (5,10,15, 20-tetrakis(4-carboxyphenyl)porphyrin, TCPP, at pH 7.0) upon the addition of methyl viologen dication (MV), as monitored over a wide MV concentration range by absorption and NMR spectra. We also characterize the effect of porphyrin concentration (Beer’s law plots) and the effect of ionic strength (phosphate buffer concentration) to sort out the factors that affect the porphyrin spectra during a typical titration experiment. Large numbers of spectral data points were taken, with each titration consisting of 30-100 individual spectra of 3200 points each. These data were analyzed both by nonlinear least squares (NLLS) at three different wavelengths and at 200 regular samplings of the full spectral data by principal component analysis (PCA) to obtain complexation constants and spectra of the complexes.
10.1021/jp0133485 CCC: $22.00 © 2002 American Chemical Society Published on Web 03/08/2002
3236 J. Phys. Chem. A, Vol. 106, No. 13, 2002 Experimental Section Materials. TCPP was purchased from Porphyrin Products (now Frontier Scientific, Inc., Logan, UT). Traces of solvents (DMF and acetic acid) were detectable by NMR and were satisfactorily removed by vacuum-drying a finely ground sample for 4 days in the dark at a pressure of 10-5 Torr with the temperature gradually raised from ambient to 100 °C. The absence of any detectable metals was verified by atomic emission spectroscopy, and the sample showed a single spot on TLC elutions. meso-Tetrakis(4-sulfonatophenyl)porphyrin, tetrasodium salt (TSPP), was purchased from Porphyrin Products and dried by the same procedure described for TCPP, except the maximum temperature was 40 °C. Methyl viologen dichloride (MV), 1,1′-dimethyl-4,4′-bipyridinium dichloride, was purchased from Pfaltz and Bauer (Waterbury, CT), treated with activated charcoal and recrystallized three times from 95% ethanol under an inert atmosphere and red light, and dried in a vacuum oven. Propyl viologen sulfonate (PVS), 1,1′-bis(3sulfonatopropyl)-4,4′-bipyridinium, was prepared by the addition of 4,4′-bipyridine, which had been recrystallized from water six times, to an excess of 1,3-propanesultone (Aldrich Chemical, Milwaukee, WI); the reaction mixture was held at 50-70 °C for 2 h, dissolved in water, precipitated with acetone, and then recrystallized and dried as described for MV.37 Cetyltrimethylammonium bromide (CTAB) was from Southwest Analytical Chemical (Austin, TX) and was recrystallized twice from ethanol. Phosphate buffer solutions were prepared from monobasic potassium phosphate, 99.999% (Aldrich), and ultrapure sodium hydroxide (Alfa Chemicals, Ward Hill, MA). Sodium chloride used to adjust ionic strength was reagent grade. Water either was distilled in a Corning apparatus or was Millipore deionized water, passed through a Millipore 0.45 µm filter just prior to use in solution preparations. UV-Vis Spectrophotometry. Spectra in the ultravioletvisible range were taken with a Shimadzu model UV260 spectrophotometer. The instrument was interfaced to a PC with UV-265 software or custom FORTRAN programs implementing GPIB bus commands. The minimum slit width (0.5 mm) and slowest scan speed were routinely used for optimum resolution. Quartz cuvettes of path lengths 0.5 mm, 1.0 mm, 1.0 cm, 2.0 cm, and 5.0 cm were used to obtain optimum spectra for the range of porphyrin concentrations studied. All spectra are reported normalized to the absorbance of a 1.0 cm path length. The temperature of the sample during the spectroscopic measurements was controlled by a Lauda K-2/RD constanttemperature bath that circulated water through an aluminum heat exchanger and sample holder that was custom designed to fit within the spectrophotometer sample compartment. The temperature was measured with a thermistor calibrated in the range 19-50 °C. NMR Spectrometry. Routine 1H NMR spectra were obtained on a Varian EM390 90 MHz spectrometer using internal TMS as reference. Studies of the complexation equilibria between TCPP and MV were performed on a 300 MHz General Electric QE-300 spectrometer in D2O solution. Stock solutions in D2O were prepared by evaporating all solvent from the corresponding aqueous stock solution, redissolving in D2O, and repeating the process shortly before the experiment (less than 4 h before all spectra were completed). The D2O solutions for NMR spectrometry were prepared from the stock solutions a few minutes before their use. Solution Preparations. Glassware and spectroscopic cuvettes were thoroughly washed with detergent, KOH in ethanol, and aqueous nitric acid, with intermediate water rinsings. Without
Clarke et al. the acid washings, traces of TCPP remained adsorbed on the glass surfaces, a general problem noted before with porphyrins.9 The glassware was next rinsed with an EDTA solution to remove trace metals, then with distilled water, then with dilute ultrapure NaOH solution to leave all acidic surface sites with sodium counterions, and finally with distilled water. Aqueous TCPP solutions were prepared from a stock 100 µM solution, which was prepared by first dissolving the requisite TCPP in a small volume of 0.10 M NaOH solution, diluting with buffer, and adjusting the pH with 0.10 M NaOH; the final pH of the stock solution was 10.2. The stock solution was stable in the dark for several months, indicated by a constant absorption spectrum of an aliquot diluted to 1.0 µM. Viologen stock solutions were prepared in distilled water and used the same day; the viologen concentration was confirmed by the ultraviolet absorption spectrum.38 Phosphate buffer solutions were prepared from ultrapure KH2PO4, adjusted to pH 7.0 with ultrapure NaOH, filtered through a 0.45 µm Millipore filter, and stored in the dark. Solution pH values were determined with an Orion model 811 pH meter. Solutions for analyses involving various concentrations of porphyrin, viologen, and buffer were either prepared individually and spectra taken immediately (always within 3 h after preparation) or monitored during a titration experiment, as described in the following section. Titration Procedures. A Gilmont model GS-3200-B ultraprecision microburet was fitted to the spectrophotometer sample compartment to allow microliter titrations with sample stirring and no intrusion in the light path. At each dispensation of titrant, stirring was halted, the buret tip was lowered just below the sample surface, titrant was dispensed, the tip was raised above the sample solution, and stirring was resumed for 1 min before a reading was taken. Typically 30-100 spectra (385-700 nm at 0.1 nm intervals) were taken during a titration, with the data transferred to a PC for analysis. Four types of titration experiments were performed: (a) variable porphyrin concentration (Beer’s law experiments) with constant buffer, (b) constant porphyrin concentration with variable salt, (c) constant porphyrin concentration with increasing viologen and constant buffer (but necessarily variable total ionic strength), (d) constant porphyrin concentration with increasing viologen but constant total ionic strength (via variable salt or buffer concentration). In the experiments with constant porphyrin and/or constant buffer, the solution in the microburet was exactly matched in concentration to the solution in the sample cuvette to avoid any dilution corrections. For example, a typical experiment of type c used a solution in the sample cuvette containing 1.0 µM TCPP in 5.0 mM phosphate buffer at pH 7.0, and the titrant solution was also 1.0 µM TCPP in 5.0 mM phosphate buffer at pH 7.0, but also contained high viologen concentration (up to 0.1 M, depending on the concentration range under study). Note that, in such experiments, the total ionic strength of the sample solution increases significantly as the addition of methyl viologen titrant increases past about 1 mM. In experiments of type d, the sample cuvette contained 1.0 µM TCPP in 62 mM phosphate buffer at pH 7.0 (ionic strength µ ) 0.15 M), and the microburet solution contained 1.0 µM TCPP in a mixture of MV and phosphate buffer that was also at an ionic strength of µ ) 0.15 M and pH 7.0. Data Analysis. The Soret regions of the TCPP spectra were decomposed into three Gaussian functions and a small linear function using the Marquardt-Levenburg algorithm.39 The linear function was used as a baseline for the entire spectrum, and absorbances were corrected by subtracting this linear component. The complexation constants and extinction coef-
Complexation Equilibria of TCPP with Viologens ficients of the complexes were evaluated from corrected absorbance data consisting of at least 30 points taken during a viologen titration experiment. Two independent methods were applied: nonlinear regression analysis for absorbances at three selected wavelengths, and principal component analysis using 200 regular samplings of the full wavelength range. The nonlinear regression analysis fit the data to a cubic equation derived from two equilibrium constants, mass balance, and total absorbance equations. PCA40-43 identified the spectrum of the 1:1 complex from the spectra observed in the two disjoint viologen concentration ranges that showed clear isosbestic points with TCPP. The two disjoint sets of two-component spectra were independently treated in the Lawton-Sylvestre technique43 whereby two allowable sectors in eigenvalue coefficient space are calculated. Since the 1:1 complex is present in both of the two-component sets, its spectral line shape was determined by locating the ray in the corresponding allowable sectors that resulted in congruent line shapes for this component. The spectral line shape for the 1:2 complex was determined by the ray on the edge of the allowable sector of eigenvalue coefficient space nearest to the highest concentration data point. PCA also verified that there were only two significant components in the isosbestic ranges and only three components in the entire range. From these data, specific concentrations of all of the components of the equilibria could be calculated throughout the concentration range and equilibrium constants evaluated. Results One of the most commonly used water-soluble anionic porphyrins is TSPP. TSPP is expected to be a tetraanion in the basic and neutral pH ranges; below pH about 4.8, protonation of TSPP is at both porphyrin nitrogens,1 rather than the sulfonates, and the resulting dianion has been studied as an interesting case of J-aggregate formation, with a unique spectroscopic signature of a sharp band at 490 nm.10,12,44,45 The related carboxy derivative TCPP is a weaker acid; no specific measurements of the pKa values for TCPP have appeared in the literature, although estimates based on the analogy to benzoic acid (pKa ) 4.2) have been occasionally assumed.6,46 Aqueous pH titrations are complicated by the very low solubility of TCPP in its un-ionized form; however, back-titration with acid of a basic aqueous solution of TCPP shows that the highest pKa value is about 6.6, with the others lying in the range 5-6. All of the studies reported here were performed in aqueous solutions of pH 7 or higher. Although we will consistently refer to the porphyrin as TCPP, without designation of the state of ionization, it is expected to be multiply ionized (primarily tetraanionic) under all the conditions used in these studies. Formation of J-aggregates from TCPP has also been suggested,45 but in these studies we never observed the characteristic spectral features that are so prominent with TSPP. Electronic Absorption Spectrum of TCPP. The UV-vis spectra of porphyrins include two major features: a series of strong visible bands (the Q bands) and an extremely intense near-ultraviolet band (the B, or Soret, band). For most free base tetraphenylporphyrins, there are four well-defined Q bands, assigned to the 0 f 0 and 0 f 1 vibronic transitions of the distinct x and y components of the lowest π f π* transition. The Soret band, assigned to the second π f π* transition, generally appears as a single intense band; however, magnetic circular dichroism studies of tetraphenylporphyrin (TPP) have shown that the Soret band also has x and y components, appearing at nearly identical positions.36 In addition, the Soret region of tetraphenylporphyrins shows a weaker shoulder at
J. Phys. Chem. A, Vol. 106, No. 13, 2002 3237
Figure 1. Soret band of 1.0 µM TCPP in 5.0 mM aqueous phosphate buffer at pH 7.0. The band is decomposed into three Gaussians and a small linear baseline.
about 400 nm, which has been considered to be the corresponding 0 f 1 transition47 or a completely separate n f π* transition.36 For the purposes of the studies reported here, which used porphyrin concentrations of 1.0 µM, the Q bands are effectively invisible, and only the Soret regions were analyzed in detail. The Soret region of the TCPP spectrum under our standard experimental conditions (1.0 µM TCPP in 5.0 mM phosphate buffer at pH 7.0) was subjected to line shape analysis. Three Gaussians and a small linear baseline were necessary for a satisfactory fit, as illustrated in Figure 1. The presence of two peaks under the main Soret band is consistent with x and y components, as assigned in the MCD spectrum of TCPP.36 Solvent and Solute Effects on the TCPP Soret Band. The overall spectrum of TCPP in its ionized form in aqueous solution at pH 7.0 is very similar to that of neutral TCPP in various organic solvents. The Q bands appear in the same relative positions and intensities (decreasing intensities at increasing wavelengths, called the etio pattern36), and the Soret region has its characteristic shape. Thus, to a first approximation, ionized TCPP in aqueous solution is considered to be a well-behaved porphyrin, whereby water solubility is simply conferred by peripheral ionic substituents that do not significantly affect the characteristic porphyrin spectroscopy. It is important to note that the class called hyperporphyrins can show relatively drastic effects on porphyrin spectra, especially with changes of ionization state.36 Hyperporphyrins involve an extension of the π conjugation beyond the porphyrin ring via quinone-like interactions with the phenyl rings, and new bands appear in the longer wavelengths, typically >700 nm. Although there are some cases of para-substituted tetraphenylporphyrins that show hyperporphyrin spectra, in particular those with strongly electrondonating substituents,48,49 it is clear that the carboxy substituent in both its ionized and un-ionized forms does not create such effects. Nevertheless, we have found that the exact location and intensity of the Soret band of TCPP varies significantly depending upon environmental conditions, in particular the solvent and the presence of other solutes (Table 1). It is important to note that all of the data in Table 1 are for highly dilute solutions where Beer’s law is obeyed, although the range in which Beer’s law holds is often rather low (Figure 2). The deviations from Beer’s law at increased concentrations have been well-documented and are generally considered to represent the formation of dimers or higher aggregates.6,7 Thus, the range
3238 J. Phys. Chem. A, Vol. 106, No. 13, 2002
Clarke et al.
TABLE 1: Solvent and Solute Effects on the Soret Absorption of TCPP solvent DMSO DMF THF 90:10 EG/H2Ob 50:50 EG/H2Ob 10:90 EG/H2Ob H2O (pH 10) H2O (pH 7) H2O (pH 11) H2O (pH 7) H2O (pH 7) H2O (pH 7)
solute
0.1 mM NaOH 5 mM bufferc 33 mM bufferd 0.5 M NaCl 2.0 M NaCl TMAPe
linear λmax (nm) regiona (×105 M-1 cm-1) 419 419 418 418 417 415 414 414.2 413 410 408 416
70 µM
8 µM 2 µM 3 µM none none
4.4 3.9 4.0 4.9 4.8 4.7 5.1 4.8 4.1
